Preparation of adipose tissue-derived cell population

Abstract

A method for preparing a stromal vascular fraction from an adipose tissue, including a step of treating an adipose tissue with an enzyme solution containing a collagenase and a neutral protease, preferably an enzyme solution free of clostripain and thermolysin, and showing not less than 1 U neutral protease activity with respect to 10,000 U collagenase activity, and recovering cells is provided by the present invention.

1 . A method for preparing a stromal vascular fraction from an adipose tissue, the method comprising the following step (1):
 (1) a step of treating an adipose tissue with an enzyme solution comprising a collagenase and a neutral protease, and showing not less than 1 U neutral protease activity with respect to 10,000 U collagenase activity, and recovering cells.   
     
     
         2 . The method according to  claim 1 , wherein the neutral protease activity of the enzyme solution is not less than 2 U with respect to 10,000 U collagenase activity. 
     
     
         3 . The method according to  claim 1 , wherein the neutral protease activity of the enzyme solution is not less than 2.5 U with respect to 10,000 U collagenase activity. 
     
     
         4 . The method according to  claim 1 , wherein the enzyme solution has a collagenase:neutral protease activity ratio of from 34,000:9 to 34,000:45. 
     
     
         5 . The method according to  claim 1 , wherein the enzyme solution has a collagenase content of not less than 500 U per 1 g of the adipose tissue. 
     
     
         6 . The method according to  claim 1 , wherein the enzyme solution has a neutral protease content of not less than 0.05 U per 1 g of the adipose tissue. 
     
     
         7 . The method according to  claim 1 , wherein the collagenase is derived from  Clostridium histolyticum.    
     
     
         8 . The method according to  claim 1 , wherein the neutral protease is derived from  Clostridium histolyticum.    
     
     
         9 . The method according to  claim 1 , wherein the enzyme solution further comprises clostripain and/or thermolysin. 
     
     
         10 . The method according to  claim 1 , wherein the enzyme solution is free of clostripain or thermolysin. 
     
     
         11 . The method according to  claim 1 , wherein the adipose tissue is a human adipose tissue. 
     
     
         12 . A method for preparing a cell population comprising a vascular endothelial cell and a vascular endothelial progenitor cell derived from an adipose tissue, the method comprising the following step (2):
 (2) a step of enriching and/or proliferating the vascular endothelial cell and the vascular endothelial progenitor cell in the stromal vascular fraction obtained by the method according to  claim 1 .   
     
     
         13 - 20 . (canceled) 
     
     
         21 . A method for dispersing adipose tissues, the method comprising the following step (1):
 (1) a step of treating an adipose tissue with an enzyme solution comprising a collagenase and a neutral protease, and showing not less than 1 U neutral protease activity with respect to 10,000 U collagenase activity.   
     
     
         22 . The method according to  claim 21 , wherein the neutral protease activity is not less than 2 U with respect to 10,000 U collagenase activity. 
     
     
         23 . The method according to  claim 21 , wherein the neutral protease activity is not less than 2.5 U with respect to 10,000 U collagenase activity. 
     
     
         24 . The method according to  claim 21 , wherein the enzyme solution has a collagenase:neutral protease activity ratio of from 34,000:9 to 34,000:45. 
     
     
         25 . The method according to  claim 21 , wherein the collagenase is derived from  Clostridium histolyticum.    
     
     
         26 . The method according to  claim 21 , wherein the neutral protease is derived from  Clostridium histolyticum.    
     
     
         27 . The method according to  claim 21 , wherein the enzyme solution further comprises clostripain and/or thermolysin. 
     
     
         28 . The method according to  claim 21 , wherein the enzyme solution is free of clostripain or thermolysin.